Instrument: Illumina HiSeq 2000
Strategy: ChIP-Seq
Source: GENOMIC
Selection: ChIP
Layout: SINGLE
Construction protocol: Prostates were dissected and then minced in 1.5ml tube with fine scissors. They were crosslinked for 15-minutes in 1% parformaldehyde. After quenching with glycine, the cross-linked chromatin was dounced and then sheared using bioruptor. Sheared chromatin was incubated with anti-rabbit IgG dynabeads pre-conjugated with 5 ug of the indicated antibody, washed, eluted, reverse cross-linked, and purified. DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 18 cycles. DNA was purified using Ampure beads and captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.